Mating type and sexual reproduction of Pseudoperonospora cubensis, the downy mildew agent of cucurbits

Abstract

The oomycete Pseudoperonospora cubensis is a leaf pathogen causing severe damage to members of the Cucurbitaceae, especially cucumber and melon. It propagates clonally by sporangia. Oospores of P. cubensis were previously observed in nature but their formation in the laboratory was never reported nor their germination or infection. Here we report on the sexual reproduction of P. cubensis under controlled conditions in the laboratory. When field isolates were inoculated singly onto detached leaves of cucurbits in growth chambers no oospores were produced. However, when pairs of selected isolates were mixed and inoculated onto detached leaves, oospores were formed in the mesophyll within 6-11 days, suggesting that P. cubensis is heterothallic, having two opposite mating types, A1 and A2. Isolates belonging to pathotype 3 were all A1 whereas isolates belonging to the new pathotype 6 were either A1 or A2. Oospores were spherical,~40 μm in diameter, hyaline to red-brown in color. Oospores were produced regularly, in large numbers, in Cucumis sativum and Cucumis melo, very seldom and in very small numbers in Cucurbita pepo, Cucurbita maxima and Citrullus lanatus, and not in Cucurbita moschata. Oospores were formed at 12.5-21°C but not at 25°C. Under moisture-saturated atmosphere oospores were also produced in leaves of intact plants. Oospores inoculated onto detached leaves in growth chambers produced F1 downy mildew lesions at 6-21 days after inoculation, many in Cucumis sativum, Cucumis melo and Cucurbita moschata, very few in Cucurbita pepo or Citrullus lanatus, and none in Cucurbita maxima. This report shows that P. cubensis is heterothallic, having A1 and A2 mating types which can cross and enable sexual reproduction in cucurbits. A preliminary report on part of the results has been published earlier.

Resurgence of cucurbit downy mildew in the United States: Insights from comparative genomic analysis of Pseudoperonospora cubensis

“…Previous studies (Cohen & Rubin, ) have suggested an association between cucurbit host types and mating types of P. cubensis . There have also been reports of both mating types being able to infect the same host species suggesting that mating type and virulence might be genetically unlinked (Cohen et al., ).…”

“…Previous studies (Cohen & Rubin, 2012) have suggested an association between cucurbit host types and mating types of P. cubensis.…”

“…Equal volumes (1:1) of sporangial suspension (2 × 10 4 spores/ml) of the tester strain and each isolate were mixed together, and the mixed suspension was then used to inoculate cantaloupe or cucumber leaves. Cantaloupe and cucumber are a favorable host substrate that supports maximum production of oospores (Cohen & Rubin, ). The first leaves of 3‐week‐old cucumber and cantaloupe leaves maintained in the glasshouse were used for inoculations.…”

“…It has been hypothesized that the resurgence of CDM in the United States was due to the introduction of a new pathotype or a new cryptic species of the pathogen (Colucci, 2008;Runge, Choi, & Thines, 2011). A new genetic recombinant from sexual reproduction has also been suspected as a possible cause for the resurgence in other parts of the world (Cohen & Rubin, 2012;Runge et al, 2011). During the past two decades, changes in the virulence structure of P. cubensis have been reported around the globe.…”

“…For example, new pathotypes have been reported in several countries (Cohen et al, 2015) and a change in population dynamics of the pathogen has been reported in the Czech Republic (Kitner et al, 2015;Lebeda et al, 2013). In addition, both mating types of P. cubensis have been found in Israel, China, the United States, and several other countries (Cohen & Rubin, 2012;Cohen et al, 2013Cohen et al, , 2014Cohen et al, , 2015Thomas, Carbone, Cohen, & Ojiambo, 2017). The global resurgence of CDM has been reviewed (e.g., Cohen et al, 2015;Holmes et al, 2015;Ojiambo et al, 2015), and understanding the genetic basis for host specialization within P. cubensis was identified as an important component in providing insights in the resurgence of CDM around the globe (Cohen et al, 2015).…”